In previous work we demonstrated that Ca spikes generated by developmentally transient Ca-dependent action potentials regulate expression of neurotransmitters. We showed that this regulation was homeostatic, such that an increase in the Ca spike activity resulted in an increase in the incidence of cells expressing inhibitory neurotransmitters (glycine and GABA) and a concomitant decrease in the incidence of excitatory neurotransmitters (glutamate and acetylcholine). Conversely, a decrease in Ca spike activity caused an increase in the incidence of excitatory neurotransmitters and a decrease in inhibitory neurotransmitters. The proposed research has two specific aims targeted at generating mutant and transgenic lines in Xenopus tropicalis useful for identifying molecules involved in the Ca spike-dependent specification of neurotransmitters and for studying activity-dependent development in general. The first specific aim uses a gain-of-function screen and the second specific aim uses gene trap insertional mutagenesis. The third specific aim characterizes a subset of the genes identified in the first and second aims. The immediate goal of this research is to identify molecules involved in Ca spike- dependent neurotransmitter specification and to generate reagents for further research in this area. In addition, we aim to generate information and mutant lines that are of value to the Xenopus community. The long-term goal is to provide information about the molecular signaling pathways that govern processes of neuronal development, which will contribute to understanding developmental disorders of the nervous system. PUBLIC HEALTH RELEVANCE: Correct specification of neurotransmitter expression during embryonic development is essential for the function of the nervous system, and both gene expression and electrical activity contribute to this process. However the mechanisms by which nature (genes) and nurture (activity) interact to specify the appropriate expression of neurotransmitters are unclear. This research will identify activity-dependent genes regulating transmitter expression. It is expected that this work will contribute to understanding developmental disorders of the nervous system in which neurotransmitter expression is altered during embryonic and post-embryonic development.